PROJECT SUMMARY Numerous studies in mice and humans have established the proximal airway basal cells (BCs) as the major multipotent stem cell population that maintains the integrity of pseudostratified epithelium through differentiation to secretory, ciliated and goblet cells. Chronic pathologies affecting the respiratory epithelium can bring about profound changes in BC biology, including BC exhaustion and dysfunction in chronic obstructive pulmonary disease (COPD) as well as emergence of proliferative BC-like populations in cystic fibrosis and cancer. Healthy luminal epithelial cells in the lung and trachea exhibit distinct apical-basal polarity, and we have found that loss this polarity stimulates signals that promote aberrant BC expansion. In particular our observations indicate that deletion of apical-localized transmembrane protein Crumbs3 leads to the dysregulation of the transcriptional regulators Yap and Taz (Yap/Taz), which have emerged as essential regulators of developmental and disease processes in the lungs and other organs. Our preliminary observations lead us to hypothesize that aberrant nuclear Yap/Taz initiate and sustain intrinsic and extrinsic signals in a microenvironment that promotes BC expansion. Analyses of polarity defective airways has revealed an interesting Yap/Taz-Neuregulin-1(Nrg1)-ERBB positive feed-back signaling cascade that we hypothesize promotes BC proliferation and self-renewal. We have also mapped notably changes in the extracellular matrix microenvironment that we hypothesize stimulates distinct Integrin-relayed signals that promote Yap/Taz activity, as well as identified a novel mesenchymal cell population that we hypothesize mediates these microenvironment changes in response to aberrant epithelial polarity. We propose that crosstalk between mesenchymal cells, extracellular matrix and the airway epithelium support aberrant BC expansion in response to polarity damage. We propose to study and target the intracellular signals mediated by Yap/Taz (AIM 1) and extracellular matrix alterations (AIM 2) that promote BC expansion, and further define how mesenchymal crosstalk contributes to phenotypes associated with epithelial polarity damage (AIM 3). Our studies will offer important molecular insight into aberrant BC expansion in airway disease, and if successful, will reveal potential biomarkers and avenues for therapeutic intervention or targeting of these poorly understood diseases, and potential new methods for expanding BC ex vivo for future regenerative therapies.